title: Not4 and Not5 modulate translation elongation by Rps7A ubiquitination, Rli1 moonlighting, and condensates that exclude eIF5A

city: Hamburg

first: Zoya

last: Ignatova

address: Martin -Luther-King-Platz 6

institute: Universitaet Hamburg

summary: Comparison of overall tRNA level and fraction of charged tRNAs of different variants

overall_design: Control cells used for tRNA charging and abundance - 2 biological replicates for each strain

first: Christine

last: Polte

first: Olesya

middle: O

last: Panasenko

first: Martine

last: Collart


title: Epigenetic Treatment of Behavioral and Physiological Deficits in a Tauopathy Mouse Model

status: Public on Aug 18 2021

city: Buffalo

first: Zhen

last: Yan

state: NY

address: 955 Main St

country: USA

institute: SUNY Buffalo

type: Expression profiling by high throughput sequencing

summary: We performed RNAseq experiments to examine genome-wide transcriptional changes in PFC of P301S Tau mice with or without UNC0642 treatment.

overall_design: mRNA via RNAseq analysis of two 6 mo. old  PS19(P301S), and two 6 mo. old control (C57BL/6 x C3H), and two 6 mo. old PS19P301S) treated with UNC0642  mice from PFC tissue were compared.

first: Jamal

last: Williams


title: IRF3-mediated pathogenicity in a murine model of human hepatitis A

city: CHAPEL HILL

first: Jason

last: Whitmire

state: NC

address: 5062 Genetic Medicine Bldg

institute: UNIVERSITY OF NORTH CAROLINA AT CHAPEL HILL

summary: Hepatitis A virus (HAV), an hepatotropic picornavirus, is a common cause of acute hepatitis in human populations. Although responsible for considerable morbidity and mortality, the mechanisms underlying HAV-mediated liver injury are poorly understood. Ifnar1-/- mice are susceptible to HAV and when infected recapitulate cardinal features of hepatitis A in humans, including serum ALT elevation, hepatocellular apoptosis, and intrahepatic inflammatory cell infiltrates. In contrast, Mavs-/- mice, while equally permissive for infection, experience no liver injury. Previous studies indicate that HAV pathogenesis in Ifnar1-/- mice is dependent upon MAVS-IRF3 signaling, but leave unresolved the role of IRF3-mediated transcription versus non-transcriptional pro-apoptotic activity of activated IRF3 in HAV-induced liver disease. Here, we compared the intrahepatic transcriptomes of HAV-infected naïve Mavs-/- and Ifnar1-/- mice using high throughput RNA sequencing, and characterized IRF3-mediated transcriptional responses associated with hepatocyte apoptosis and liver inflammation.

overall_design: Liver mRNA profiles of Ifnar1-/- ,Mavs-/- and Irf3S1/S1Ifnar1-/- mice 7 days or 14 days after HAV infection

first: Stanley

last: Lemon

first: You

last: Li


title: A cis-acting mechanism mediates trancriptional memory at Polycomb target genes in mammals

status: Public on Aug 19 2021

city: Paris

first: Daniel

last: Holoch

state: Cedex 05

address: 26 rue d'Ulm

department: Genetics and Developmental Biology Unit

country: France

institute: Institut Curie, Centre de Recherche

type: Genome binding/occupancy profiling by high throughput sequencing

type: Methylation profiling by high throughput sequencing

summary: This SuperSeries is composed of the SubSeries listed below.

overall_design: Refer to individual Series


title: The RNA helicase Dbp7 promotes domain V/VI compaction and stabilization of inter-domain interactions during early 60S assembly

city: Göttingen

first: Katherine

last: Bohnsack

address: Humboldtallee 23

country: Germany

institute: University Medical Centre Göttingen

summary: Early pre-60S ribosomal particles are poorly characterized, highly dynamic complexes that undergo extensive rRNA folding and compaction concomitant with assembly of ribosomal proteins and exchange of assembly factors. Pre-60S particles contain numerous RNA helicases, which are likely regulators of accurate and efficient formation of appropriate rRNA structures. Here we reveal binding of the RNA helicase Dbp7 to domain V/VI of early pre-60S particles in yeast and show that in the absence of this protein, dissociation of the Npa1 scaffolding complex, release of the snR190 folding chaperone, recruitment of the A3 cluster factors and binding of the ribosomal protein uL3 are impaired. uL3 is critical for formation of the peptidyltransferase center and is responsible for stabilizing interactions between the 5’ and 3’ ends of the 25S, an essential pre-requisite for subsequent pre-60S maturation events. Highlighting the importance of pre-ribosome remodeling by Dbp7, our data suggest that in the absence of Dbp7 or its catalytic activity, early pre-ribosomal particles are targeted for degradation.

overall_design: Wild-type BY4741 yeast cells or cells expressing genomically His-Tev-ProtA-tagged Dbp7 were grown in the presence of 4-thiouracil, crosslinked using light at 365 nm and analysed by CRAC.

first: Gerald

middle: R

last: Aquino

first: Philipp

last: Hackert

first: Nicolai

last: Krogh

first: Kuan-Ting

last: Pan

first: Mariam

last: Jaafar

first: Anthony

middle: K

last: Henras

first: Henrik

last: Nielsen

first: Henning

last: Urlaub

middle: E

first: Markus

middle: T


title: Creld2 function during unfolded protein response is essential for liver metabolism homeostasis

city: Bonn

first: Elvira

last: Mass

address: Carl-Troll str

institute: LIMES

summary: The unfolded protein response (UPR) is associated with the metabolic function of the liver, yet it is not well understood how endoplasmic reticulum (ER) disturbance might influence metabolic homeostasis. Here, we describe the physiological function of the Cysteine-rich with EGF-like domains 2 (Creld2), which has been previously characterized as a downstream target of the ER-stress signal transducer Atf6. Creld2 enhances protein folding and degradation through its interaction with proteins involved in UPR, thereby, promoting tolerance of chronic stress and recovery from acute stress. Using a genetic knockout model, we show that Creld2-deficiency leads to a dysregulated UPR, and causes the development of hepatic steatosis in male mice, while females are protected. We observed this sex dimorphism also in humans with fatty liver disease, with only males showing an accumulation of CRELD2 protein in the liver. These results reveal a Creld2 function at the intersection between UPR and metabolic homeostasis and suggest a mechanism in which chronic ER stress underlies fatty liver disease in males.

overall_design: Analysis of liver RNA expression of mice from two different in vivo models for non-alcoholic fatty liver induction. Dietary in vivo model mice were fed either control diet (CD) or high fat diet (HFD) for 12 weeks, or HFD for 8 weeks followed by CD for 4 weeks (HFD>CD). Pharmacological in vivo model mice were administered either a sucrose control, thapsigargin (TG, 0.5 mg/kg) or tunicamycin (TM, 1mg/kg) via intraperitoneal injection and sacrificed after 48 hours.

first: Paul

last: Kern

first: Nora

last: Balzer

first: Franziska

last: Bender

first: Alex

last: Frolov

first: Jan-Peter

last: Sowa

first: Thomas

last: Ulas

first: Lorenzo

last: Bonaguro

first: Özlem

last: Kücükoglu

first: Christoph

last: Thiele

first: Joachim

middle: L

last: Schultze

first: Ali

last: Canbay

first: Reinhard

last: Bauer


title: Disrupted Circadian Oscillations in Type 2 Diabetes are Linked to Altered Rhythmic Mitochondrial Metabolism in Skeletal Muscle [Affymetrix]

status: Public on Aug 17 2021

city: Copenhagen

last: Altintas

address: Blegdamsvej 3B

department: NNF Center for Basic Metabolic Research

country: Denmark

institute: Copenhagen University

summary: Circadian rhythms are generated by an auto-regulatory feedback loop composed of transcriptional activators and repressors. Disruption of circadian rhythms contributes to Type 2 diabetes (T2D) pathogenesis. We elucidated whether altered circadian rhythmicity of clock genes is associated with metabolic dysfunction in T2D. Transcriptional cycling of core clock genes BMAL1, CLOCK, and PER3 was altered in skeletal muscle from individuals with T2D and this was coupled with reduced number and amplitude of cycling genes and disturbed circadian oxygen consumption. Inner-mitochondria associated genes were enriched for rhythmic peaks in NGT, but not T2D, and positively correlated with insulin sensitivity. ChIP-sequencing identified CLOCK and BMAL1 binding to inner-mitochondrial genes associated with insulin sensitivity, implicating regulation by the core clock. Inner-mitochondria disruption altered core-clock gene expression and free-radical production, phenomena that were restored by resveratrol treatment. We identify bi-directional communication between mitochondrial function and rhythmic gene expression, processes which are disturbed in diabetes.

overall_design: Skeletal muscle biopsies were obtained from men with either T2D or normal glucose tolerance (NGT). Primary myotubes cultures were prepared, synchronized and harvested every 6 hours over 42 hours and the transcriptome was analyzed by RNA-sequencing.

first: Brendan

last: Gabriel

last: Altıntaş

first: Mikael

last: Rydén

first: Hui

last: Gao

first: Romain

last: Barrès

first: Anna

last: Krook

first: Juleen

last: Zierath


title: A microRNA signature for early detection and RNA therapy in Alzheimer´s disease

city: Goettingen

first: Md Rezaul

last: Islam

address: von siebold str. 3a

institute: DZNE

type: Non-coding RNA profiling by high throughput sequencing

summary: Since changes in cognitive function develop slowly over time, Alzheimer’s disease patients are often diagnosed at an advanced stage of molecular pathology, a time point when causative treatments fail. Thus, there is great need for inexpensive and minimal invasive biomarker that could be used in screening approaches with the aim to identify individuals at risk, that can then undergo further diagnostics and eventually stratified therapies. In this study we use an integrative approach combining the analysis of human data and model systems to identify a circulating 3-microRNA signature that reflect key processes linked to neural homeostasis and informs about cognitive decline. We furthermore provide evidence that expression changes of this signature represent relevant patho-mechanisms in the brain and are a suitable target for RNA therapeutics towards dementia.

overall_design:
Longitudinal analysis of smallRNAseq data along aging in mice in learning group
After habituation at 12 months, cognitive performance was assessed in C57BL/6 mice at age of 13.5, 15 and 16.5 months. Blood was collected from retroorbital sinus at given time points and isolated RNA was subjected to small RNA sequencing.

first: Islam

last: Rezaul

first: Fischer

last: Andre


title: Identification of circulating miRNA molecular signature for erectile dysfunction in type 2 diabetes

city: Harbin

first: Ping

last: Wang

state: Heilongjiang

address: Harbin Medical University

country: China

institute: Harbin Medical University

summary: we apply miRNA sequencing from blood samples of 10 DMED patients and 10 DM controls to study the mechanism of miRNAs action on DMED.

first: Hong

last: Qiao

first: Haibo

last: Xu


title: Epigenetic therapies in ovarian cancer alter repetitive element expression in a TP53-dependent manner: ChIPseq

city: Washington

first: James

last: McDonald

state: DC

address: 800 22nd St, NW, Suite 8960

institute: The George Washington University

summary:
Abstract: Epithelial ovarian carcinomas (OC) are particularly deadly due to intratumoral heterogeneity, resistance to standard-of-care therapies, and poor response to alternative treatments such as immunotherapy. Targeting the OC epigenome with DNA methyltransferase inhibitors (DNMTi) or histone deacetylase inhibitors (HDACi) increases immune signaling and recruits CD8+ T cells and NK cells to fight OC in murine models. This increased immune activity is caused by increased transcription of repetitive elements (RE) that form double-stranded RNA (dsRNA) and trigger an interferon response. To understand which REs are affected by epigenetic therapies in OC, we assessed the effect of DNMTi and HDACi on OC cell lines and patient samples. Subfamily-level (TEtranscripts) and individual locus-level (Telescope) analysis of REs showed that DNMTi treatment upregulated more REs than HDACi treatment. Upregulated REs were predominantly LTR and SINE subfamilies, and SINEs exhibited the greatest loss of DNA methylation upon DNMTi treatment. Cell lines with TP53 mutations exhibited significantly fewer upregulated REs with epigenetic therapy than wild type TP53 cell lines. This observation was validated using isogenic cell lines; the TP53 mutant cell line had significantly higher baseline expression of REs but upregulated fewer upon epigenetic treatment. In addition, p53 activation increased expression of REs in wild type but not mutant cell lines. These data give a comprehensive, genome-wide picture of RE chromatin and transcription-related changes in OC after epigenetic treatment and implicate p53 in RE transcriptional regulation.
The data in this data series represent the analysis of P53 ChIPseq libraries generated from the A2780 ovarian carcinoma cell lines. Each cell line was treated with Nutlin-3A or 5-azacytidine. Each library was sequenced, trimmed, aligned, and analyzed with MACS2 and IDR. The ChIP-seq peaks were used to analyze how P53 regulated repetitive element transcription in response to epigenetic therapies.

overall_design: Two replicates of P53 ChIP-seq were performed in A2780 cell lines given mock, Nutlin-3A, or 5-azacytidine. A matching single replicate input sample was made for each treatment. The two pulldown replicates were analyzed by IDR in comparison to the single input replicate.

middle: I

first: Noor

last: Diab

first: Elisa

last: Arthofer

first: Melissa

last: Hadley

first: Tomas

last: Kanholm

first: Uzma

last: Rentia

first: Stephanie

last: Gomez

first: Angela

last: Yu

first: Erin

last: Grundy

first: Olivia

last: Cox

first: Michael

middle: J

last: Topper

first: Xiaoyun

last: Xing

first: Pamela

last: Strissel

first: Reiner

last: Strick

first: Ting

first: Stephen

middle: B

last: Baylin

first: Katherine

last: Chiappinelli


title: Epigenetic therapies in ovarian cancer alter repetitive element expression in a TP53-dependent manner: ATACseq

first: James

summary:
Abstract: Epithelial ovarian carcinomas (OC) are particularly deadly due to intratumoral heterogeneity, resistance to standard-of-care therapies, and poor response to alternative treatments such as immunotherapy. Targeting the OC epigenome with DNA methyltransferase inhibitors (DNMTi) or histone deacetylase inhibitors (HDACi) increases immune signaling and recruits CD8+ T cells and NK cells to fight OC in murine models. This increased immune activity is caused by increased transcription of repetitive elements (RE) that form double-stranded RNA (dsRNA) and trigger an interferon response. To understand which REs are affected by epigenetic therapies in OC, we assessed the effect of DNMTi and HDACi on OC cell lines and patient samples. Subfamily-level (TEtranscripts) and individual locus-level (Telescope) analysis of REs showed that DNMTi treatment upregulated more REs than HDACi treatment. Upregulated REs were predominantly LTR and SINE subfamilies, and SINEs exhibited the greatest loss of DNA methylation upon DNMTi treatment. Cell lines with TP53 mutations exhibited significantly fewer upregulated REs with epigenetic therapy than wild type TP53 cell lines. This observation was validated using isogenic cell lines; the TP53 mutant cell line had significantly higher baseline expression of REs but upregulated fewer upon epigenetic treatment. In addition, p53 activation increased expression of REs in wild type but not mutant cell lines. These data give a comprehensive, genome-wide picture of RE chromatin and transcription-related changes in OC after epigenetic treatment and implicate p53 in RE transcriptional regulation.
The data in this data series represent the analysis of ATAC-seq libraries generated from the A2780, Hey, Kuramochi, and TykNu ovarian carcinoma cell lines. Each cell line was treated with ITF-2357 (HDACi), 5-azacytidine (DNMTi), or a combination of both. ATAC-seq libraries were sequenced, trimmed, aligned, and analyzed with the HMMRATAC software.

overall_design: Single replicates of ATAC-seq were performed in A2780, Hey, Kuramochi, and TykNu cell lines given mock, ITF-2357, 5-azacytidine, or ITF-2357/5-azacytidine combined.

last: Wang


title: Fasting Mimicking Diet cycles prevent effects of a high fat diet on cardiometabolic risk factors and lifespan

city: Los Angeles

first: Alice

last: Mouton

state: CA

address: 610 Charles E. Young Dr. East

department: Ecology and Evolutionary Biology

institute: UCLA

summary: Diet-induced obesity is the central cause of diabetes, cardiovascular disease as well as metabolic syndrome. Here, we have studied the efficacy of cycles of a 4-5 day fasting mimicking diet (FMD) in inhibiting high-fat, high calorie diet (HFCD) -induced obesity in mature female C57BL/6 mice. We show that a monthly 5-day cycle of FMD inhibit HFCD-mediated obesity by causing a reduction in calorie intake and accumulation of visceral and subcutaneous fat depots without lean body mass loss. FMD cycles also increase cardiac vascularity, function and stress resistance, and reverse the hypercholesterolemia caused by the HFCD. The sustained activation of adipocyte genes associated with mitochondrial metabolism and biogenesis and the sustained ketogenesis in the HFCD-fed mice subjected to monthly cycles of FMD indicate a reprogramming of fat cell metabolism that is likely to be at the center of obesity reversal. All these improvements could explain the protection from early mortality elicited by the high-fat, high calorie diet.

overall_design: 18 C57BL/6 mice were divided into 3 groups and were fed ad libitum either a commercial high fat and high calorie diet (HFCD) (60% calories from fat) or a control diet (10% calories from fat), or a HFCD diet combined once every 4 weeks with cycles of a 5 day FMD (HFCD+FMD). Starting at cycle 6, the 5 day FMD was followed by 2 days of a control diet before the switch to HFCD. Heart and adipose tissue were harvested after 11 cycles of FMD, 4 days after the end of FMD cycle. RNA samples were extracted using the RNeasy Mini Kit following instructions (Qiagen). Libraries were built using the KAPA RNA Hyper+RiboErase HMR kit (Roche) and sequenced on an Illumina HiSeq3000 machine.

first: Amrendra

last: Mishra


title: Epigenetic therapies in ovarian cancer alter repetitive element expression in a TP53-dependent manner: Isogenic TP53 Cell Line RNA-seq

last: McDonald

summary:
Abstract: Epithelial ovarian carcinomas (OC) are particularly deadly due to intratumoral heterogeneity, resistance to standard-of-care therapies, and poor response to alternative treatments such as immunotherapy. Targeting the OC epigenome with DNA methyltransferase inhibitors (DNMTi) or histone deacetylase inhibitors (HDACi) increases immune signaling and recruits CD8+ T cells and NK cells to fight OC in murine models. This increased immune activity is caused by increased transcription of repetitive elements (RE) that form double-stranded RNA (dsRNA) and trigger an interferon response. To understand which REs are affected by epigenetic therapies in OC, we assessed the effect of DNMTi and HDACi on OC cell lines and patient samples. Subfamily-level (TEtranscripts) and individual locus-level (Telescope) analysis of REs showed that DNMTi treatment upregulated more REs than HDACi treatment. Upregulated REs were predominantly LTR and SINE subfamilies, and SINEs exhibited the greatest loss of DNA methylation upon DNMTi treatment. Cell lines with TP53 mutations exhibited significantly fewer upregulated REs with epigenetic therapy than wild type TP53 cell lines. This observation was validated using isogenic cell lines; the TP53 mutant cell line had significantly higher baseline expression of REs but upregulated fewer upon epigenetic treatment. In addition, p53 activation increased expression of REs in wild type but not mutant cell lines. These data give a comprehensive, genome-wide picture of RE chromatin and transcription-related changes in OC after epigenetic treatment and implicate p53 in RE transcriptional regulation.
The data in this data series represent the analysis of triplicate RNAseq libraries generated from two isogenic lines derived from the Hey ovarian carcinoma cell line. HC2 is TP53 wild type as is the parent Hey line. HH23 was edited to express TP53 with the R175H mutation. Each line was treated with 5-azacytidine, Nutlin-3A, or a combination of both. RNA-seq libraries were sequenced, trimmed, aligned, and analyzed with TEtranscripts and Telescope software. Note that because the core facility did not reach our target number of reads after the first run, a second run was performed on the same library sample. These two runs were treated as one sample in the analysis.

overall_design: Three biological replicates of RNA-seq were performed in isogenic Hey-derived cell lines designated HC2 (TP53 WT) and HH23 (TP53 Mutant R175H) given mock, Nutlin-3A (N3A), 5-azacytidine (AZA), or combined N3A/AZA.

first: Noor

last: Diab

first: Elisa

last: Arthofer

first: Melissa


title: A microRNA signature for early detection and RNA therapy in Alzheimer´s disease (3-microRNA mimic)

first: Md Rezaul

last: Islam

overall_design: Analysis of RNAseq data after treatment of brain organoids with lipid nanoparticles packaged with either 3-microRNA mimic cocktail or negative control siRNA

first: Rezaul

first: Andre

last: Fischer


title: A microRNA signature for early detection and RNA therapy in Alzheimer´s disease (small RNA)

overall_design: Longitudinal analysis of smallRNAseq data along aging in mice


title: Mucosa samples from patients with Familial Adenomatous Polyposis (FAP)

city: Houston

first: Eduardo

last: Vilar-Sanchez

state: Texas

address: 1515 Holcombe Blvd, Unit 1360

institute: UT MD Anderson Cancer Center

summary: The genetic alterations of intestinal carcinogensis have been observed in multiple comparative studies. However,  transcriptomic characteristics of at-risk muocsa and adenomas from patients with FAP are still lacking. Therefore, our aim was to characterize the transcriptomic profile of at-risk mucosa of FAP patients.

overall_design: 10 samples were analyzed using Illumina HiSeq

last: Vilar


title: A20 deficiency in myeloid cells protects mice from diet-induced obesity and insulin resistance due to increased fatty acid metabolism

city: Zwijnaarde

first: Liesbet

last: Martens

address: Technologiepark 927

department: VIB Inflammation Research Center

country: Belgium

institute: VIB-University of Ghent

summary: Obesity-induced inflammation is a major driving force in the development of insulin resistance, type 2 diabetes (T2D) and related metabolic disorders. During obesity, macrophages accumulate in the visceral adipose tissue, creating a low grade inflammatory environment through the production of inflammatory cytokines. NF-κB signaling is a central coordinator of these inflammatory responses and is tightly regulated by the anti-inflammatory protein A20. To understand the role of inflammatory macrophages in insulin resistance and T2D development, we fed myeloid-specific A20 deficient mice (A20myel-KO mice) with a high fat diet, and demonstrate that these mice are completely protected from diet-induced obesity and insulin resistance, despite an inflammatory environment in their metabolic tissues. Macrophages lacking A20 show impaired mitochondrial respiratory function and metabolize more palmitate both in vitro and in vivo. Given the increased macrophage numbers in A20myel-KO mice, we hypothesize that A20 deficient macrophages rely more on palmitate oxidation and metabolize the fat present in the diet, resulting in a lean phenotype and protection from metabolic disease. These findings reveal an unexpected role for A20 in regulating macrophage immunometabolism.

overall_design: Bulk RNA sequencing of FACS-purified CD206+ and CD11c+ adipose tissue macrophage subclasses isolated from epididymal adipose tissue (eWAT) of A20myel-KO and wild-type C57BL/6J mice.

first: Leen

last: Catrysse

first: Bastiaan

last: Maes

first: Parul

last: Mehrotra

first: Arne

first: Esther

last: Hoste

first: Christian

last: Maueröder

first: Anneleen

last: Remmerie

first: Anna

last: Bujko

first: Karolina

last: Slowicka

first: FMozes

last: Sze

first: Hanna

last: Vikkula

first: Bart

last: Ghesquière

first: Charlotte

last: Scott

first: Yvan

last: Saeys

last: van de Sluis

first: Kodi

last: Ravichandran

first: Sophie

last: Janssens

first: Geert

last: van Loo


title: mRNA-seq read counts of peripheral blood mononuclear cells from congenital generalized lipodystrophy patients and their gender/aged-matched controls

city: Taipei City

first: Yen-Hua

last: Huang

address: No. 155, Sec. 2, Li-Nong St., Beitou Dist

department: Institute of Biomedical Informatics

country: Taiwan

institute: National Yang-Ming University

summary: Congenital generalized lipodystrophy (CGL) is an autosomal recessive disorder characterized by defective adipose tissue, extreme insulin resistance, and early onset of diabetes. There are four types of congenital generalized lipodystrophy based on the causative genetic alterations. The symptoms and the degrees of disease progression are varied among all affected individuals, which might be due to unknown genetic modifiers. To identify potential predictive biomarkers associated with the disease progressions, we recruited 7 patients (cgl2_p1,cgl2_p2,cgl2_p3,cgl2_p4,cgl2_p5,cgl2_p6,cgl2_p7),as well as gender/aged-matched controls (cgl2_c1,cgl2_c2,cgl2_c3,cgl2_c4,cgl2_c5,cgl2_c6,cgl2_c7).The total RNA samples were extracted from the participants' peripheral blood mononuclear cells, followed by RNA-Seq. To find the molecular signatures that might be associated with disease progression, patients were further categorized into sub-groups based on the results of biochemical analysis and their clinical symptoms (e.g. diabetes, metabolic syndrome and metal retardation). DEGs were identified for each sub-groups of patients, and pathway analysis was performed to explore the underlying dysregulated mechanisms.

overall_design: PBMC from 7 patients and 7 gender/aged-matched controls were extracted total RNA, followed by RNA-Seq.

first: YH

first: TC

last: Su

first: CH

last: Wang

first: SL

last: Wong

last: Chien

first: YT

first: WL

last: Hwu

first: NC

last: Lee


title: A cis-acting mechanism mediates trancriptional memory at Polycomb target genes in mammals [RNA-Seq]

first: Daniel

last: Holoch

summary: Epigenetic inheritance of gene expression states enables a single genome to maintain distinct cellular identities. How histone modifications contribute to this process remains unclear. Using global chromatin perturbations and local, time-controlled modulation of transcription, we establish the existence of epigenetic memory of transcriptional activation for genes that can be silenced by the Polycomb group. This property emerges during cell differentiation and allows genes to be stably switched following a transient transcriptional stimulus. This transcriptional memory state at Polycomb targets operates in cis; however, rather than relying solely on read-and-write propagation of histone modifications, the memory is also linked to the strength of activating input opposing Polycomb proteins and therefore varies with the cellular context. Our data and computational simulations suggest a model whereby transcriptional memory arises from double-negative feedback between Polycomb-mediated silencing and active transcription. Epigenetic memory at Polycomb targets thus depends not only on histone modifications but also on the gene-regulatory network and underlying identity of a cell.

overall_design:
Transcriptomic analysis (RNA-seq) of mouse neural progenitor cells in untreated (2 replicates), PRC2-inhibitor-treated (2 replicates) and PRC2-inhibitor-washout (2 replicates) conditions (6 samples).
Transcriptomic analysis (RNA-seq) of mouse embryonic stem cells in untreated (2 replicates), PRC2-inhibitor-treated (2 replicates) and PRC2-inhibitor-washout (2 replicates) conditions (6 samples).
Transcriptomic analysis (RNA-seq) of immortalized mouse embryonic fibroblast line A (= C2) in wild-type (2 replicates), Ezh2 KO (2 replicates) and Ezh2 rescue (2 replicates) conditions (6 samples).
Transcriptomic analysis (RNA-seq) of immortalized mouse embryonic fibroblast line B (= C26) in wild-type (2 replicates), Ezh2 KO (2 replicates) and Ezh2 rescue (2 replicates) conditions (6 samples).
Nascent RNA analysis (chromosome-associated-RNA-seq) of immortalized mouse embryonic fibroblast line A (= C2) in wild-type (2 replicates), Ezh2 KO (2 replicates) and Ezh2 rescue (2 replicates) conditions (6 samples).
Nascent RNA analysis (chromosome-associated-RNA-seq) of immortalized mouse embryonic fibroblast line B (= C26) in wild-type (2 replicates), Ezh2 KO (2 replicates) and Ezh2 rescue (2 replicates) conditions (6 samples).
Transcriptomic analysis (RNA-seq) of immortalized mouse embryonic fibroblast line B bearing the DD-dCas9-VPR transgene and either sgRNAs targeting the Nr2f1 promoter, either before induction (2 replicates) or more than 9 days after the conclusion of a transient induction (2 replicates), or sgRNAs targeting the Foxa1 promoter, either before induction (2 replicates) or more than 9 days after the conclusion of a transient induction (2 replicates) (8 samples).
Targeted analysis (RNA-seq) of Nr2f1 mRNA levels in an equal mixture of RNA from wild-type immortalized mouse embryonic fibroblast line B cells in which Nr2f1 expression has been induced and RNA from Nr2f1-mutant cells in which Nr2f1 expression is repressed, or in an equal mixture of RNA from wild-type and Nr2f1-mutant cells in which Nr2f1 expression has been induced, or in RNA from each of two heterokaryon clones resulting from fusion of wild-type cells in which Nr2f1 expression has been induced and RNA from Nr2f1-mutant cells in which Nr2f1 expression is repressed, or in RNA from each of these two clones after dCas9-VPR-mediated induction of Nr2f1 expression, or in RNA from each of these two clones more than 9 days after the conclusion of such an induction, and targeted analysis of Foxa1 mRNA in a parallel series of conditions (16 samples).

first: Michel

last: Wassef

first: Cecilia

last: Lövkvist

first: Dina

last: Zielinski

first: Setareh

last: Aflaki

first: Bérangère

last: Lombard

first: Tiphaine

last: Héry

first: Damarys

last: Loew

first: Martin

last: Howard

first: Raphaël

last: Margueron


title: Donor bone marrow-derived macrophage MHC II drives neuroinflammation and cognitive deficits during chronic GVHD in mice

city: Brisbane

first: Dylan

middle: Patrick

last: Carter-Cusack

state: Queensland

address: 300 Herston Rd

department: Department of Immunology

country: Australia

institute: QIMR Berghofer Medical Research Institute

summary: Graft-versus-host disease (GVHD) remains the leading cause of non-relapse mortality after allogeneic stem cell transplantation for haematological malignancies. Manifestations of GVHD in the central nervous system (CNS) present as neurocognitive dysfunction in up to 60% of patients, however, the mechanisms driving chronic GVHD in the CNS are yet to be elucidated. Our studies of murine chronic GVHD revealed a spatial learning and memory deficit associated with inflammation in the hippocampus underlain by persistent Ifng upregulation. By flow cytometry, we observed a proportional shift in the donor-derived T-cell population in the chronic GVHD brain from early CD8 dominance to later CD4 sequestration. Hippocampal RNA sequencing identified perturbations to structural and functional synapse-related gene expression, together with the upregulation of genes associated with IFN-γ responses and antigen presentation. Neuroinflammation in the cortex of mice and humans during acute GVHD was recently shown to be mediated by resident microglia-derived TNF. In contrast, infiltration of pro-inflammatory MHC class II+ donor bone marrow-derived macrophages (BMDM) was identified as a distinguishing feature of CNS chronic GVHD. Donor BMDM, which comprised up to 50% of the CNS myeloid population, exhibited a transcriptional signature distinct from resident microglia. Recipients of MHC class II knockout bone marrow grafts exhibited attenuated neuroinflammation, suggestive of a critical role for donor BMDM in chronic CNS GVHD. Our identification of disease mediators distinct from those in the acute phase indicates the necessity to pursue alternative therapeutic targets for late-stage neurological manifestations.

overall_design: Experiment 1: Hippocampi from GVHD and non-GVHD mice (day 80 post transplant) were isolated and harvested for RNA isolation (n = 3 GVHD, 3 TCD)           Experiment 2: Donor and host microglia isolated from GVHD mice and host microglia isolated from TCD mice (day 70 post transplant) were isolated via FACS and then harvested for RNA isolation (n = 6 GVHD Donor, 4 GVHD Host, 3 TCD Host)

first: Rachael

middle: C

last: Adams

first: Samreen

middle: N

last: Shaik

first: Genesis

last: Llanes

first: Rebecca

last: Johnston

first: Gregory

last: Quaife-Ryan

first: Glen

last: Boyle

first: Lambros

last: Koufariotis

first: Andreas

last: Möller

first: Marc

last: Ruteinberg

first: Bruce

last: Blazar

first: Jana

last: Vukovic

first: Kelli

middle: P

last: MacDonald


title: A microRNA signature for early detection and RNA therapy in Alzheimer´s disease (miR-181a-5p)

first: Md Rezaul

last: Islam

overall_design: Analysis of RNAseq data from primary hippocampal neurons treated with lipid nanoparticles packaged with either miR-181a-5p mimic or negative control siRNA

first: Rezaul

first: Andre

last: Fischer


title: A cis-acting mechanism mediates trancriptional memory at Polycomb target genes in mammals [ChIP-Seq]

first: Daniel

last: Holoch

overall_design:
H3K27me3 enrichment profiles (CUT&RUN-seq) of mouse neural progenitor cells in untreated (2 replicates), PRC2-inhibitor-treated (2 replicates) and PRC2-inhibitor-washout (2 replicates) conditions (6 samples).
H3K27me3 enrichment profiles (ChIP-seq) of immortalized mouse embryonic fibroblast line A (= C2) in wild-type (2 replicates), Ezh2 KO (2 replicates) and Ezh2 rescue (2 replicates) conditions (6 samples).
H3K27me3 enrichment profiles (ChIP-seq) of immortalized mouse embryonic fibroblast line B (= C26) in wild-type (2 replicates), Ezh2 KO (2 replicates), Ezh2 rescue (2 replicates) conditions (6 samples).
Input sample for ChIP-seq of immortalized mouse embryonic fibroblast line A (= C2) (1 sample).
RNA polymerase II enrichment profiles (ChIP-seq) of immortalized mouse embryonic fibroblast line A (= C2) in Ezh2 KO (2 replicates) (2 samples).
H3K4me3 enrichment profiles (ChIP-seq) of immortalized mouse embryonic fibroblast line A (= C2) in Ezh2 KO (2 replicates) (2 samples).
H3K36me3 enrichment profiles (ChIP-seq) of immortalized mouse embryonic fibroblast line A (= C2) in Ezh2 KO (2 replicates) (2 samples).

first: Michel

last: Wassef

first: Cecilia

last: Lövkvist

first: Dina

last: Zielinski

first: Setareh

last: Aflaki

first: Bérangère

last: Lombard

first: Tiphaine

last: Héry

first: Damarys

last: Loew

first: Martin

last: Howard

first: Raphaël


title: Epigenetic therapies in ovarian cancer alter repetitive element expression in a TP53-dependent manner: TykNu Tiplicate RNA-seq

first: James

last: McDonald

summary:
Abstract: Epithelial ovarian carcinomas (OC) are particularly deadly due to intratumoral heterogeneity, resistance to standard-of-care therapies, and poor response to alternative treatments such as immunotherapy. Targeting the OC epigenome with DNA methyltransferase inhibitors (DNMTi) or histone deacetylase inhibitors (HDACi) increases immune signaling and recruits CD8+ T cells and NK cells to fight OC in murine models. This increased immune activity is caused by increased transcription of repetitive elements (RE) that form double-stranded RNA (dsRNA) and trigger an interferon response. To understand which REs are affected by epigenetic therapies in OC, we assessed the effect of DNMTi and HDACi on OC cell lines and patient samples. Subfamily-level (TEtranscripts) and individual locus-level (Telescope) analysis of REs showed that DNMTi treatment upregulated more REs than HDACi treatment. Upregulated REs were predominantly LTR and SINE subfamilies, and SINEs exhibited the greatest loss of DNA methylation upon DNMTi treatment. Cell lines with TP53 mutations exhibited significantly fewer upregulated REs with epigenetic therapy than wild type TP53 cell lines. This observation was validated using isogenic cell lines; the TP53 mutant cell line had significantly higher baseline expression of REs but upregulated fewer upon epigenetic treatment. In addition, p53 activation increased expression of REs in wild type but not mutant cell lines. These data give a comprehensive, genome-wide picture of RE chromatin and transcription-related changes in OC after epigenetic treatment and implicate p53 in RE transcriptional regulation.
The data in this data series represent the analysis of triplicate RNAseq libraries generated from the TykNu ovarian carcinoma cell line treated with ITF-2357 (HDACi), 5-azacytidine (DNMTi), or a combination of both. RNA-seq libraries were sequenced, trimmed, aligned, and analyzed with TEtranscripts and Telescope software.

overall_design: Three biological replicates of RNA-seq were performed in the TykNu cell line given mock, ITF-2357, 5-azacytidine, or combined ITF-2357/5-azacytidine.

first: Noor

last: Diab

first: Elisa

last: Arthofer

first: Melissa

last: Hadley

first: Tomas

last: Kanholm

first: Uzma

last: Rentia

first: Stephanie

last: Gomez

first: Angela

last: Yu

first: Erin

last: Grundy

first: Olivia

last: Cox

first: Michael

last: Topper

first: Xiaoyun

last: Xing

first: Pamela

last: Strissel

first: Reiner

last: Strick

first: Ting

last: Wang

first: Stephen

last: Baylin

first: Katherine

last: Chiappinelli


title: Enteric pathogens induce tissue tolerance and prevent neuronal loss from subsequent infections

city: New York

first: Tomasz

last: Ahrends

address: 1230 York Ave

institute: The Rockefellere University

summary: The enteric nervous system (ENS) controls several intestinal functions including motility and nutrient handling, which can be disrupted by infection-induced neuropathies or neuronal cell death. We investigated possible tolerance mechanisms preventing neuronal loss and disruption in gut motility after pathogen exposure. We found that following enteric infections, muscularis macrophages (MMs) acquire a tissue-protective phenotype that prevents neuronal loss and dysmotility during subsequent challenge with unrelated pathogens. Bacteria-induced neuroprotection relied on activation of gut-projecting sympathetic neurons and signaling via b2-adrenergic receptors (b2AR) on MMs. In contrast, helminth-mediated neuroprotection was dependent on T cells and systemic production of interleukin (IL)-4 and -13 by eosinophils, which induced arginase-expressing MMs that prevented neuronal loss from an unrelated infection located in a different intestinal region. Collectively, these data suggest that distinct enteric pathogens trigger a state of disease- or tissue tolerance that preserves ENS number and functionality.

overall_design: C57BL/6J mice were s.c. injected with water (non-infected, NI) or infected with spiB, Sv or Yp. Ileum MMs were sorted for RNA-seq.


title: Microencapsulated 3D co-cultures recapitulate phenotypic features of B lymphomas

status: Public on Aug 16 2021

city: Rennes

first: Julien

last: Saint-Vanne

address: 2 avenue du professeur Léon Bernard - CS 34317

summary: Non-Hodgkin B-cell lymphomas (B-NHL) mainly develop within lymph nodes (LN) as densely packed aggregates of tumor cells and their surrounding microenvironment, creating a tumor niche specific to each lymphoma subtypes. In vitro preclinical models mimicking biomechanical forces, cellular microenvironment, and 3D organization of B-cell lymphomas remain scarce, while all these parameters constitute key determinants of lymphomagenesis and drug resistance. Using a microfluidic method based on cell encapsulation inside permeable, elastic, and hollow alginate microspheres, we developed a new tunable 3D-model incorporating lymphoma B cells, extracellular matrix and/or stromal cells.Using RNA-sequencing, we reported that, in contrast to classical 2D cocultures, our 3D lymphoma model initiated a coevolution of these two cell types, recapitulating the physiopathology of B lymphoma tissue.

overall_design: DOHH2 and HLY1 cell lines were cultured in 2D or 3D conditions (alginate spheroids) with or without tonsil stromal cells (TSC) before cell sorting of CD20pos B cells and RNAseq

first: Claire

last: Lamaison

first: Laurence

last: Bepoldin

first: Karin

last: Tarte

first: Frédéric

last: Mourcin

last: Saint Vanne

first: Valérie

last: Morvan


title: Analysis of MGE transcriptomes with or without Ctnnb1 knockout in mouse through RNA sequencing

city: Shanghai

first: yanhua

last: du

address: 227 South Chongqing Road, Huangpu District

department: Shanghai Institute of Immunology

institute: Shanghai Jiaotong University School of Medicine

summary:
Purpose: Understand the cues that orchestrate the expansion or differentiation of medial ganglionic eminence (MGE) progenitors no matter in vivo or in vitro.
Methods: Total RNA from each sample was used to prepare the library. Then the libraries were sequenced at 50bp single read on an Illumina HiSeq 2500 platform. Sequencing reads from each sample were mapped to the mouse reference genomes mm10 by using TopHat v2.1.1. The mapped reads were further analyzed by cufflinks v1.3.0 and the expression levels for each transcript were quantified as Fragments Per Kilobase of transcript per Million mapped reads (FPKM). For differential expression analysis, sequencing counts at the gene level were obtained using HTSeq v 0.9.1. R package DESeq2 was then used to identify differential expressed genes between different conditions.
Results: Transcriptome profiling reveals that ablation of β-catenin in MGE cells leads to advanced neuronal differentiation, while activation of Wnt/β-catenin signaling keeps the MGE cells in an undifferentiated progenitor state.
Conclusions: Wnt signaling is a key player in governing self-renewal vs. terminal differentiation of MGE progenitors both in vivo and in vitro.

overall_design: mRNA profiles of MGE cells (E14.5) with or without Ctnnb1 knockout were generated by RNA sequencing, using Illumina Hiseq 2500 platform.

first: Lin

last: Ma

first: Yanhua

last: Du


title: A novel therapeutic strategy targeting the mesenchymal phenotype of malignant pleural mesothelioma by suppressing LSD1 [ATAC-seq]

city: Bunkyo-Ku

first: Ken

last: Tajima

state: Tokyo

address: 2-1-1 Hongo

department: Department of Respiratory Medicine

country: Japan

institute: Juntendo University

summary: ATAC-seq analysis of following knock down of LSD1 in ACC-MESO-1 cells.

overall_design: We performed ATAC-seq analysis in ACC-MESO-1 cell infected with shLSD1 or control.

first: Aditya

last: Wirawan


title: A microRNA signature for early detection and RNA therapy in Alzheimer´s disease (miR-146a)

first: Md Rezaul

last: Islam

overall_design: Analysis of RNAseq data from immortalized microglial cells treated with lipid nanoparticles packaged with either miR-146a mimic or negative control siRNA

first: Rezaul

first: Andre

last: Fischer


title: A microRNA signature for early detection and RNA therapy in Alzheimer´s disease (miR-mix)

overall_design: Analysis of RNAseq data from 7 months old APPS1-21 mice and littermate controls treated with lipid nanoparticles packaged with either anti-miR-mix or negative control siRNA


title: A microRNA signature for early detection and RNA therapy in Alzheimer´s disease (anti-miR-mix)

overall_design: Analysis of RNAseq data from old and young mice treated with lipid nanoparticles packaged with either anti-miR-mix or negative control siRNA


title: The RNA helicase Dbp7 promotes domain VI compaction and stabilization of inter-domain interactions during early 60S assembly

first: Nicolai

last: Krogh

department: Department of Cellular and Molecular Medicine

institute: University of Copenhagen

summary: Early pre-60S ribosomal particles are poorly characterized, highly dynamic complexes that undergo extensive rRNA folding and compaction concomitant with assembly of ribosomal proteins and exchange of assembly factors. Pre-60S particles contain numerous RNA helicases, which are likely regulators of accurate and efficient formation of appropriate rRNA structures. Here we reveal binding of the RNA helicase Dbp7 to domain V/VI of early pre-60S particles in yeast and show that in the absence of this protein, dissociation of the Npa1 scaffolding complex, release of the snR190 folding chaperone, recruitment of the A3 cluster factors and binding of the ribosomal protein uL3 are impaired. uL3 is critical for formation of the peptidyltransferase center and is responsible for stabilizing interactions between the 5’ and 3’ ends of the 25S, an essential pre-requisite for subsequent pre-60S maturation events. Highlighting the importance of pre-ribosome remodeling by Dbp7, our data suggest that in the absence of Dbp7 or its catalytic activity, early pre-ribosomal particles are targeted for degradation.

overall_design: All samples were analyzed in biological triplicates.

first: Gerald

last: Aquino

first: Philipp

last: Hackert

first: Kuan-Ting

last: Pan

first: Mariam

last: Jaafar

first: Anthony

last: Henras

first: Henrik

last: Nielsen

first: Henning

last: Urlaub

last: Bohnsack

first: Markus


title: Metronomic Radiotherapy Reverses Tumor Immune Desertification and Resistance to Immunotherapy

city: Epalinges

first: Raphael

last: Genolet

address: Biopole 3, Chemin des Boveresses 155

department: Departement d'oncologie UNIL-CHUV

country: Switzerland

institute: Centre Hospitalier Universitaire Vaudois (CHUV)

summary: We report major tumor regressions in a subset of advanced cancer patients for which cold tumors were treated with low-dose radiotherapy, cyclophosphamide, aspirin, and ICB (NCT03728179). Unbiased analyses of biopsies revealed T-cell infiltration, up-regulation of type I IFN, and Th1 signatures as well as  down-regulation of M2 macrophage and epithelial to mesenchymal transition gene-signatures, and a more oligoclonal TCR repertoire after radio-combinatorial immunotherapy, in responding patients.

overall_design: Evaluation of blood TCR repertoire at different time point

first: Frenanda

middle: G

last: Herrera

first: Catherine

last: Ronet

first: Maria

last: Ochoa de Olza

first: George

last: Coukos


title: Gene expression profile of HSC/Ps and mature myeloid cells in MDS mutant clone

city: Hachioji

first: Yoshihiro

last: Hayashi

address: 1432-1 Horinouchi

institute: Tokyo University of Pharmacy and Life Sciences

summary: MDS are characterized by bone marrow (BM) failure due to ineffective hematopoiesis. However, the underlying mechanisms of ineffective hematopoiesis, in which clonal expansion coexists with accelerated cell death, remain to be elucidated. Recently, we detected CBL exon 8/9 deletion mutation (CBLΔE8/9) in several MDS patients with RUNX1 mutations. The CBLΔE8/9/RUNX1S291fs mice (CR-mice) developed a variety of MDS phenotypes, such as pancytopenia, dysplasia, and ineffective hematopoiesis in BM. RNA-Seq of HSC/progenitor (HSC/P) and mutant mature myeloid BM cells revealed that gene signature induced in the CR-mice is similar to that in MDS patients with BM failure. Especially, innate immune-related genes were significantly dysregulated.

overall_design: Expression profiles of mRNA in HSC/Ps and Ly6G+ myeloid cells from CBL/RUNX1 mutants mice and their control mice.

first: Hironori

last: Harada


city: Lausanne

first: Isaac

last: Crespo

state: Vaud

address: Rue de Bugnon

department: CHUV-Oncology

institute: UNIL

summary: The development of approaches for inflaming cold tumors is critical for increasing response to immunotherapy (IMT) reliant upon the re-invigoration of T cells for tumor control. Here we report, in a pre-clinical advanced ID8 ovarian cancer model, that metronomic low-dose radiotherapy (LDRT; 1 Gy) promotes T-cell infiltration and enables responsiveness to treatment including low-dose cyclophosphamide (CP), and combinatorial IMT comprising immune checkpoint blockade (ICB) targeting PD-1 and CTLA-4, and anti-CD40 agonist antibody, for significant tumor control and survival. Responses to this radio-combinatorial immunotherapy (RACIM) relied upon re-programming of both innate and adaptive immunity. We further identified RACIM-induced intratumoral dendritic cell states with co-stimulatory capacity and expressing the stress ligand RAE1, as well as of polyfunctional NKG2D+TCF1-PD1+CD4+T cells. We further report major tumor regressions in a subset of advanced cancer patients for which cold tumors were treated with LDRT, CP, aspirin, and ICB (NCT03728179). Unbiased analyses of biopsies revealed T-cell infiltration, up-regulation of type I IFN, and Th1 signatures as well as  down-regulation of M2 macrophage and epithelial to mesenchymal transition gene-signatures, and a more oligoclonal TCR repertoire after RACIM, in responding patients.

overall_design: Evaluation of the impact on immune cells of low dose whole abdominal radiotherapy  in orthotopic ovarian cancer. In this experiments, mice were injected ID8 ovarian cancer cells (5 million). 21 days later mice were treated: 1) control/NT, cyclophophamide(CP)+ aPD1+aCTLA4+aCD40ag (IT), or CP + IT + low-dose Rx (1 gray each) 8 days later, a second round. 5 days later mice were sacrificed and tumors processed.

first: Fernanda

first: Melita

last: Irving


title: A microRNA signature for early detection and RNA therapy in Alzheimer´s disease (miR-148a-3p)

first: Md Rezaul

last: Islam

summary: Since changes in cognitive function develop slowly over time, Alzheimer’s disease patients are often diagnosed at an advanced stage of molecular pathology, a time point when causative treatments fail. Thus, there is great need for inexpensive and minimal invasive biomarker that could be used in screening approaches with the aim to identify individuals at risk, that can then undergo further diagnostics and eventually stratified therapies. In this study we use an integrative approach combining the analysis of human data and model systems to identify a circulating 3-microRNA signature that reflect key processes linked to neural homeostasis and informs about cognitive decline. We furthermore provide evidence that expression changes of this signature represent relevant patho-mechanisms in the brain and are a suitable target for RNA therapeutics towards dementia.

overall_design: Analysis of RNAseq data from primary hippocampal neurons treated with lipid nanoparticles packaged with either miR-148a-3p mimic or negative control siRNA

first: Rezaul

first: Andre

last: Fischer


title: Exome sequencing of KP and KPM lung tumors.

status: Public on Aug 15 2021

first: Nadine

last: Fournier

address: Agora Cancer Research Center, Bugon 25A

institute: Agora Cancer Research Center & Swiss Institute of Bioinformatics

summary: Immune checkpoint blockade (ICB) with PD1 or PDL1 antibodies has been approved for the treatment of non-small cell lung cancer (NSCLC). However, only a minority of the patients respond and sustained remissions are rare. Both chemotherapy and anti-angiogenic drugs may improve tumor response to ICB in mouse models and patients with cancer. Here, we used genetically engineered mouse models of KrasG12D/p53null NSCLC, including a mismatch repair-deficient variant (KrasG12D/p53null/Msh2null) with higher mutational burden, and longitudinal imaging to study tumor response and resistance to combinations of ICB, anti-angiogenic therapy, and chemotherapy. Anti-angiogenic blockade of VEGFA and angiopoietin-2 markedly slowed progression of established tumors but, contrary to findings in other mouse cancer models, addition of a PD1 or PDL1 antibody was not beneficial and even accelerated progression of some of the tumors. We found that anti-angiogenic treatment facilitated tumor infiltration by PD1+ regulatory T cells (T-regs), which were more efficiently targeted by the PD1 antibody than CD8+ T cells. Both tumor-associated macrophages (TAMs) of bone-marrow origin, which are CSF1R-dependent, and TAMs of alveolar origin, which are sensitive to cisplatin, contributed to establishing a TGFB-rich tumor microenvironment that supported PD1+ T-regs. Dual TAM targeting with a combination of cisplatin and a CSF1R inhibitor fully abated T-regs, redirected the PD1 antibody to CD8+ T cells, and improved the efficacy of anti-angiogenic immunotherapy, achieving regression of the majority of the tumors.

overall_design: Exome sequencing of KP (KrasLSL-G12D/+; p53fl/fl) and KPM (KrasLSL-G12D/+; p53fl/fl;Msh2–/–) lung tumors.

first: Amaia

last: Martinez-Usatorre

last: Zangger

first: Michele

last: De Palma


title: Overcoming microenvironmental resistance to anti-PD-1 immunotherapy in lung cancer models

summary: This SuperSeries is composed of the SubSeries listed below.

overall_design: Refer to individual Series


title: Single-cell RNA sequencing reveal pre-existing and persistent subpopulations of cells associated with relapse of prostate cancer [scRNA-seq 2]

city: London

first: Paolo

last: Cremaschi

state: Non-US

address: 72 Huntley Street

department: Oncology

country: United Kingdom

institute: UCL Cancer Institute

summary: Prostate cancer is heterogeneous and patients would benefit from methods that stratify clinically indolent from more aggressive forms of the disease. We employed single-cell RNA sequencing in models of early treatment response and resistance to enzalutamide. In doing so, we identified pre-existing and treatment-persistent cell subpopulations that possess transcriptional stem-like features and regenerative potential when subjected to treatment. We found distinct chromatin landscapes associated with enzalutamide treatment and resistance that are linked to alternative transcriptional programs. Transcriptional profiles characteristic of persistent stem-like cells were able to stratify the treatment response of patients. Ultimately, we show that defining changes in chromatin and gene expression in single-cell populations from pre-clinical models can reveal hitherto unrecognized molecular predictors of treatment response. This suggests that high analytical resolution of pre-clinical models may powerfully inform clinical decision-making.

overall_design: Single-cell RNA sequencing of LNCaP cells treated with 10µM enzalutamide for 168 hours, VCaP parental cells (treated with DMSO for 48 hours), and VCaP cells treated with 10µM enzalutamide for 48 hours.

first: Karolina

last: Nowakowska

first: Sinja

last: Taavitsainen

first: Daniel

last: Wetterskog

first: Alfonso

last: Urbanucci

first: Gerhardt

last: Attard


title: Gene expression profile of HSC/Ps and mature myeloid cells in Mdivi-1 treated MDS mutant clone

first: Yoshihiro

summary: MDS are characterized by bone marrow (BM) failure due to ineffective hematopoiesis. However, the underlying mechanisms of ineffective hematopoiesis, in which clonal expansion coexists with accelerated cell death, remain to be elucidated. Recently, we detected CBL exon 8/9 deletion mutation (CBLΔE8/9) in several MDS patients with RUNX1 mutations. The CBLΔE8/9/RUNX1S291fs mice (CR-mice) developed a variety of MDS phenotypes, such as pancytopenia, dysplasia, and ineffective hematopoiesis in BM. RNA-Seq of HSC/progenitor (HSC/P) and mutant mature myeloid BM cells revealed that gene signature induced in the CR-mice is similar to that in MDS patients with BM failure. Especially, inflammation and innate immune-related genes were significantly dysregulated. Mdivi-1 treatment significantly attenuated inflammaotry signaling activation and rescued MDS phenotypes.

overall_design: Expression profiles of mRNA in HSC/Ps and Ly6G+ myeloid cells from CBL/RUNX1 mutants mice with or without Mdivi-1 treatment.


title: Expression profile comparison of enriched bone marrow-derived tumor associated macrophages, tumor-associated alveolar macrophages, CD4 T cells, and total tumor-derived cells in lung cancer models with or without anti-PD1 and anti-VEGFA/ANGPT2 treatments.

overall_design: Expression profile comparison of enriched bone marrow-derived tumor associated macrophages (BM-TAMs), tumor-associated alveolar macrphages (T-AM), CD4 T cells, and total (unfractioned) tumor-derived viable cells, from KrasG12D/p53null/Msh2null NSCLC tumors of mice untreated (IgG), treated with anti-VEGFA/ANGPT2 antibody (A2V), or A2V + anti-PD1 antibodies.


title: A novel therapeutic strategy targeting the mesenchymal phenotype of malignant pleural mesothelioma by suppressing LSD1

first: Ken

last: Tajima


title: The effect of stress on the transcriptomes of circulating immune cells in patients with Gulf War Illness

city: Fort Lauderdale

first: Lubov

last: Nathanson

state: Florida

address: 3321 College Ave.

department: INIM

institute: Nova Southeastern University

summary:
Aims: In an effort to gain further insight into the underlying mechanisms tied to disease onset and progression of Gulf War Illness (GWI), our team evaluated GWI patient response to stress utilizing RNA-Seq.
Main Methods: The protocol included blood collection before (baseline), during (peak of exercise, measured by VO2 max), and after exercise challenge (recovery – four hours post-exercise challenge). Peripheral blood mononuclear cell (PBMC) transcriptomics data were analyzed using DESeq2 and DEXseq software to understand why GWI patients process stressors differently from their healthy counterparts.
Key findings: Our findings validate previously identified dysregulation of immune and inflammatory pathways among GWI patients as well as highlight novel immune and inflammatory markers of disease activity. These results provide a foundation for future research efforts in understanding GWI pathophysiology and creating targeted treatments.
Significance: Gulf War Illness is a complex, chronic, and debilitating multi-system illness impacting 25%-30% of the U.S. troops deployed to the 1990-1991 Gulf War. The condition is characterized by medically unexplained fatigue and affects multiple organ systems. Because the underlying mechanisms are largely unknown, patients receive symptom-based treatment, rather than targeting fundamental biological processes. To the best of our knowledge, this is the first study that applies RNA-Seq to analyze the effect of GWI, and the response to stressors in GWI, on the transcriptomic changes in circulating immune cells.

overall_design: RNA-seq of total RNA from peripheral blood mononuclear cells of GWI patients and matched healthy controls that underwent the exercise challenge. Blood was drawn at T0 (baseline, before exercise), T1 (VO2max, peak of exercise) and T2 (recovery, four hours after T1) time points.

first: Derek

last: Van Booven

first: Oskar

last: Zarnowski

first: Melanie

last: Perez

first: Leonor

last: Sarria

first: Fanny

last: Collado

first: Kyle

last: Hansotia

first: Sean

last: Riegle

first: Tali

last: Finger

first: Mary

middle: A

last: Fletcher

first: Nancy

last: Klimas


title: Hobit identifies TRM precursors that are regulated by Eomes

city: Amsterdam

first: Klaas

last: van Gisbergen

state: Select one

address: Plesmanlaan 125

department: Department of Hematopoiesis

country: Netherlands

institute: Sanquin Research and Landsteiner Laboratory

summary: Tissue-resident memory CD8+ T cells (TRM) constitute a non-circulating memory T cell subset that provides early protection against re-infection. However, how TRM arise from antigen-triggered T cells has remained unclear. Exploiting the TRM-restricted expression of Hobit, we developed TRM reporter/deleter mice to study TRM differentiation. We found that Hobit was upregulated in a subset of LCMV-specific T cells located within peripheral tissues during the effector phase of the immune response. These Hobit+ effector T cells were identified as TRM precursors, given that their depletion substantially decreased TRM development, but not the formation of circulating memory T cells. Adoptive transfer experiments of Hobit+ effector T cells corroborated their biased contribution to the TRM lineage. Transcriptional profiling of Hobit+ effector T cells underlined the early establishment of TRM properties including downregulation of tissue exit receptors and upregulation of TRM-associated molecules. Importantly, we identified Eomes as a key factor instructing the early bifurcation of circulating and resident lineages. These findings establish that commitment of TRM occurs early in antigen-driven T cell differentiation and reveal the molecular mechanisms underlying this differentiation pathway.

overall_design: We used RNA-seq to analyse FACS-sorted effector CD8+ Tcells. A total of 45 tissue samples where analysed; 15 conditions with 3 biological replicates. Lymphocytes were extracted from Liver, kidney, spleen and SI-IEL tissues from TRM reporter/deleter mice after LCMV infection. Naïve T-cells were extracted from the spleen as reference.

first: Loreto

last: Parga-Vidal

first: Felix

middle: M

last: Behr

first: Natasja

last: Kragten

first: Benjamin

last: Nota

first: Thomas

middle: H

last: Wesselink

first: Inga

last: Kavazović

first: Laura

last: Covill

first: Margo

last: Schuller

first: Yenan

last: Bryceson

last: Wensveen

first: Rene

last: van Lier

first: Teunis

last: van Dam

first: Regina

last: Stark


title: Elevated N-linked glycosylation of IgG variable regions in myasthenia gravis disease subtypes

status: Public on Aug 13 2021

city: New Haven

first: Steven

last: Kleinstein

state: Ct

address: 300 George St.

department: Pathology

institute: Yale University

summary: Elevated N-linked glycosylation of immunoglobulin G variable regions (IgG-VN-Glyc) is an emerging molecular phenotype associated with autoimmune disorders. To test the broader specificity of elevated IgG-VN-Glyc, we studied patients with distinct subtypes of myasthenia gravis (MG), a B cell-mediated autoimmune disease. Our experimental design included adaptive immune receptor repertoire sequencing to quantify and characterize N-glycosylation sites in the global B cell receptor repertoire, proteomics to examine glycosylation patterns of the circulating IgG, and production of human-derived recombinant autoantibodies, which were studied with mass spectrometry and antigen binding assays to confirm occupation of glycosylation sites and determine whether they alter binding. We found that the frequency of IgG-VN-Glyc motifs was increased in the B cell repertoire of MG patients when compared to healthy donors. Motifs were introduced by both biased V gene segment usage and somatic hypermutation. IgG-VN-Glyc could be observed in the circulating IgG in a subset of MG patients. Autoantigen binding, by patient-derived MG autoantigen-specific monoclonal antibodies with experimentally confirmed presence of IgG-VN-Glyc, was not altered by the glycosylation. Our findings extend prior work on patterns of variable region N-linked glycosylation in autoimmunity to MG subtypes. Although occupied IgG-VN-Glyc motifs are found on MG autoantigen-specific monoclonal antibodies, they are not required for binding to the autoantigen in this disease.

overall_design: A MuSK MG cohort (N=3) was defined using BCR repertoire sequencing derived from a previous study. Another n = 11 AChR MG and n = 9 heathy control subjects were selected for BCR based adaptive immune receptor repertoire-sequencing or AIRR-Seq using PBMC derived RNA for this study. In total, serum samples from all 3 MuSK MG subjects and 9 AChR MG subjects (8 overlapping with paired AIRR-Seq) cohort were also investigated for the presence of VH gene glycosylation-specific signatures in the serum. For patients with MuSK MG diagnoses and patient AChR MG-1 with an AChR MG diagnosis, longitudinal serum samples were collected. A patient with Rheumatoid Arthritis (RA) (n = 1) was enrolled in a research study at University of California San Francisco (UCSF) for pathogen and autoantibody detection.

first: Caleigh

last: Mandel-Brehm

first: Miriam

last: Fichtner

first: Ruoyi

last: Jiang

first: Valerie

last: Winton

first: Sara

last: Vazquez

first: Minh

last: Pham

first: Kenneth

last: Hoehn

first: Neil

last: Kelleher

first: Richard

last: Nowak

first: Michael

last: Wilson

first: Joseph

last: DeRisi

first: Kevin

last: O’Connor


title: Single-cell landscape of peripheral immune responses to fatal SFTS

status: Public on Aug 14 2021

city: Beijing

first: Wei

last: Liu

address: 20 Dong-Da Street, Fengtai District

institute: Beijing Institute of Microbiology and Epidemiology, State Key Laboratory of Pathogen and Biosecurity

summary: We performed scRNA-seq on 27 PBMC samples from 15 SFTS patients (including four deceased and 11 surviving) and four healthy controls. We found that the immune cell landscape was reprogrammed in SFTS and characterized by monocyte shift to intermediate type along with complement activation, perturbation of plasmablasts composition, and highly exhausted T cells, all were correlated with lethal consequences. We identified the overexpression of interferon-stimulated genes across most immune cell types post SFTSV infection, which were simultaneously related to older age, high viremia and hyper-inflammation. These data collectively supported the intermediate monocytes and IFN-I-inducible plasmablasts to be major targets for SFTSV infection, and proposed the pivotal role of IFN-I response in exacerbating hyper-inflammation and lethal SFTS, which might help to identify therapeutic strategies for SFTS.

overall_design: A cohort study was used to identify the role of peripheral immune responses in SFTS infection. A total of 15 SFTS patients, including 11 with the CT values of RT-qPCR ranging from 25 to 30 and four with the CT values of RT-qPCR less than 25, were prospectively sampled for scRNA-seq and flow cytometry analysis.

first: Hao

last: Li

first: Xiaokun

first: Shouming

last: Lv

first: Xuefang

last: Peng

first: Ning

last: Cui

first: Tong

last: Yang

first: Zhendong

first: Chun

last: Yuan

first: Jiaying

last: Yao

first: Zan

first: Xiaolei

last: Ye

first: Xiaoai

last: Zhang

first: Shu

last: Zhu


title: Dietary suppression of MHC-II expression in intestinal epithelial cells enhances intestinal tumorigenesis

city: Berkeley

first: Karthik

last: Shekhar

address: 101D Gilman Hall

institute: University of California, Berkeley

summary: Little is known about how interactions between diet, intestinal stem cells (ISCs) and immune cells impact the early steps of intestinal tumorigenesis. Here, we show that a high fat diet (HFD) reduces the expression of the major histocompatibility complex II (MHC-II) genes in intestinal epithelial cells including ISCs. This decline in epithelial MHC-II expression in a HFD correlates with reduced diversity of the intestinal microbiome and is recapitulated in antibiotic treated and germ-free mice on a control diet. Mechanistically, pattern recognition receptor (PRR) and IFN g signaling regulate epithelial MHC-II expression where genetic ablation of these signaling pathways dampen MHC-II epithelial expression. Interestingly,  upon loss of the tumor suppressor gene Apc in a HFD, MHC-II-  ISCs harbor greater in vivo tumor-initiating capacity than their MHC-II+ counterparts when transplanted into immune-component hosts but not immune-deficient hosts, thus implicating a role for epithelial MHC-II-mediated immune surveillance in suppressing tumorigenesis. Finally, ISC-specific genetic ablation of MHC-II in engineered Apc-mediated intestinal tumor models increases tumor burden in a cell autonomous manner. These findings highlight how a HFD perturbs a microbiome – stem cell – immune cell crosstalk in the intestine and contributes to tumor initiation through the dampening of MHC-II expression in pre-malignant ISCs.

overall_design: single-cell RNA-seq of Lgr5+ intestinal stem cells derived from control or HFD mice

first: Semir

last: Beyaz

first: Omer

last: Yilmaz

first: Mose

last: Biton


title: PARP1-mediated PARylation activity is essential for oligodendroglial differentiation and CNS myelination

city: Sacramento

first: Fuzheng

last: Guo

address: 2425 Stockton Blvd

institute: University of California Davis

summary: The function of poly(ADP-ribosyl) polymerase 1 (PARP1) in myelination and remyelination of the central nervous system (CNS) remains enigmatic. Here we report that PARP1 is an intrinsic driver for oligodendroglial development and myelination. Genetic PARP1 depletion impairs the differentiation of oligodendrocyte progenitor cells (OPCs) into oligodendrocytes and impedes CNS myelination. Mechanistically, PARP1-mediated PARylation activity is not only necessary but also sufficient for OPC differentiation. At the molecular level, we identify the RNA-binding protein Myef2 as a PARylated target which controls OPC differentiation through PARylation-modulated de-repression of myelin protein expression. Furthermore, PARP1’s enzymatic activity is necessary for oligodendrocyte and myelin regeneration after demyelination. Together, our findings suggest that PARP1-mediated PARylation activity may be a potential therapeutic target for promoting OPC differentiation and remyelination in neurological disorders characterized by arrested OPC differentiation and remyelination failure such as multiple sclerosis.

overall_design: Three Parp1 cKO mice and three littermate control mice (both sexes) were  injected with tamoxifen at P1, P2, and P3 and the brain was harvested for total RNA extraction and NGS RNS-sequencing. Downstream analysis was followed to assess the gene expression level across the 6 samples.

first: Yan

last: Wang


title: DSCR1 upregulation enhances dural meningeal lymphatic drainage to attenuate amyloid pathology of Alzheimer’s disease

city: Ulsan city

first: Chiyeol

last: Choi

state: 울산

address: Republic of Korea

country: South Korea

institute: UNIST

summary: Purpose: To further understand the beneficial effects of DSCR1 on the lymphatic system of 5XFAD mice, we investigated the transcriptional landscape of meningeal lymphatic endothelial cells (mLECs) by performing RNA sequencing.

overall_design: mRNA profiles of meningeal lymphatic endothelial cells (mLECs) were isolated from 3-4 mice of each strain using MACS, in quadruplicate

first: Jiyoung

last: Park


title: scRNA-seq analysis of pharmacological HCN channel block with ZD7288 on rat primary cortical neuronal stem cells

city: Cologne

first: Malte

last: Stockebrand

address: Kerpener Str. 62

institute: DZNE e.V.


title: scRNA-seq analysis (well-based) of pharmacological HCN channel block with ZD7288 on rat primary cortical neuronal stem cells

summary: We analysed transcriptomic changes of single cell transcriptomes from primary rat cortical neuronal stem cells (NSC) after pharmacologically blocking HCN channels with ZD7288 compared to untreated NSC. This well-based scRNA-seq aproach revealed a pronounced cell cycle arrest in G1 phase of ZD7288 treated (HCN blocked) NSC.

overall_design: scRNA-seq (well based) of two groups of neuronal stem cells, 4 replicates each group with a total of 500 cells: 200 treated 300 non-treated cells with addition of 5 wells containing bulk RNA (positive control) or 5 empty wells (negative control)

first: Dirk

last: Isbrandt

first: Anna

last: Schlusche


title: Identification of LZTFL1 as a candidate effector gene at a COVID-19 risk locus (ATAC-seq)

city: Headington

first: Damien

last: Downes

state: Oxfordshire

address: John Radcliffe Hospital

department: MRC Weatherall Institute of Medicine

institute: The University of Oxford

summary: To identify regions of open chromatin ATAC-seq was carried out in Blood outgrowth endothelial cells (BOECs) and NCI-H441 Epithelial cells

overall_design: ATAC-seq was carried out in BOECs (n=4) and H441 Epithelial cells (n=4).


title: RNA-seq analysis of helios deficient hematopoietic stem and progenitor cells.

city: illkirch

first: giovanni

last: cova

address: 1 Rue Laurent Fries

institute: igbmc

summary: Here we addressed the mRNA transcriptome of Long-Term Hematopoietic Stem Cells (LT-HSC) and Multipotent Progenitor (MPP) -3 and -4 purified from WT and Helios KO mice. By analyzing the differentially expressed genes among WT and KO condition we found that Helios deletion affects transcription mostly in the LT-HSC compartment and favor a megakaryocyte mRNA lineage priming at the expense of the lymphoid one.

overall_design: Long-term hematopoietic stem cells (LT-HSC) and multipotent progenitors myeloid and lymphoid biased (MPP-3 and -4 respectively) isolated from the bone marrow from 3 WT and 3 Helios KO mice by flow cytometry. Total RNA was extracted from LT-HSC, MPP3 and MPP4 cells and used to prepared libraries that were sequenced. The analysis of the differentially expressed genes in WT and KO cell populations revealed that Helios deletion favor a megakaryocyte lineage priming mRNA profiling at the expense of the lymphoid one in these cells.

first: Giovanni

last: Cova

first: Peggy

last: Kirstetter

first: Philippe

last: Kastner

first: Susan

last: Chan


title: ChIP-seq HELIOS (Ikzf2)

summary: Here we addressed the genome-wide DNA binding profile of Helios transcription factor in early hematopoietic progenitor cells. We reported Helios DNA target sites by performing Chromatin Immuno-Precipitation (ChIP) assay on genomic DNA from the early hematopoietic progenitor cells line HPC7.

overall_design: We sequenced the HPC7 chromatin immunoprecipitated with Helios as well as the Input chromatin.

first: Marie-Celine

last: Deau


title: Transcriptome analysis of Filifactor alocis in co-culture with Porphyromonas gingivalis

city: Loma Linda

first: Yuetan

last: Dou

address: 11021 Campus Street AH114

department: Microbiology and Molecular Genetics

institute: Loma Linda University

summary: We use high through put RNA sequenceing technology to study the genome-wide expression profile in an oral pathogen Filifactor alocis when co-cultured with another key-stone oral pathogen Porphyromonas gingivalis under anaerobic conditions and oxidative stress conditions.

overall_design: Comparison of F. alocis for co-culture with P. gingivalis under anaerobic conditions and oxidative stress conditions.

first: Hansel

last: Fletcher


title: scRNA-seq analysis (drop-seq) of pharmacological HCN channel block with ZD7288 on rat primary cortical neuronal stem cells

summary: We analysed transcriptomic changes of single cell transcriptomes from primary rat cortical neuronal stem cells (NSC) after pharmacologically blocking HCN channels with ZD7288 compared to untreated NSC. This drop-seq scRNA-seq aproach showed accumulation of ZD7288 treated cells in G1 phase and confirmed our previous findings from a well-based scRNA-seq approach with cell cycle arrest in G1 after HCN channel block.

overall_design: scRNA-seq of two neuronal stem cell populations, one treated one non-treated


title: Identification of LZTFL1 as a candidate effector gene at a COVID-19 risk locus (ChIP-seq)

summary: To identify regulation by the CEBPB transcription factor in epithelial, fibroblast and endothelial cells ChIP-seq was carried out for both the transcription factor and marks of active transcription (H3K27ac).

overall_design: ChIP-seq was carried out for one replicate each of H441 lung epithelial cells, IMR-90 liung fibroblast cells, human umbilical vein endothelial cells (HUVEC) and blood outgrowth endothelial cells (BOEC).


title: Characterization of immune cells infiltrating islets using scRNA-seq

city: MILWAUKEE

first: Yi-Guang

last: Chen

state: WI

address: 8701 Watertown Plank Road

institute: Medical College of Wisconsin

summary: In type 1 diabetes (T1D) autoreactive CD8 T cells infiltrate pancreatic islets and destroy insulin-producing β cells. Progression to T1D onset is a chronic process, which suggests that the effector activity of β-cell autoreactive CD8 T cells needs to be maintained throughout the course of disease development. The mechanism that sustains diabetogenic CD8 T cell effectors during the course of T1D progression has not been completely defined. Here we used single-cell RNA sequencing to gain further insight into the phenotypic complexity of islet-infiltrating CD8 T cells in NOD mice. We identified two functionally distinct subsets of activated CD8 T cells, CD44highTCF1+CXCR6- and CD44highTCF1-CXCR6+, in islets of prediabetic NOD mice. Compared to CD44highTCF1+CXCR6- CD8 T cells, the CD44highTCF1-CXCR6+ subset expressed higher levels of inhibitory and cytotoxic molecules and was more prone to apoptosis. Adoptive cell transfer experiments revealed that CD44highTCF1+CXCR6- CD8 T cells, through continuous generation of the CD44highTCF1-CXCR6+ subset, were more capable than the latter population to promote insulitis and the development of T1D. We further showed that direct interleukin-27 (IL-27) signaling in CD8 T cells promoted the generation of terminal effectors from the CD44highTCF1+CXCR6- population. These results indicate that islet CD44highTCF1+CXCR6- CD8 T cells are a progenitor-like subset with self-renewing capacity and under an IL-27 controlled mechanism they differentiate into the CD44highTCF1-CXCR6+ terminal effector population. Our study provides new insight into the sustainability of the CD8 T cell response in the pathogenesis of T1D.

overall_design: Gene expression profiles of single immune cells infiltrating islets of 7-week old and 12-week old NOD mice prepared using 10x Genomics Chromium platform

first: Ashley

last: Ciecko

first: David

last: Schauder


title: Prenatal overexpression of platelet-derived growth factor receptor A results in central nervous system hypomyelination

status: Public on Aug 12 2021

city: New York

first: Oren

middle: Josh

last: Becher

state: NY

address: 1468 Madison Avenue, A21-02

department: Pediatrics

institute: Mount Sinai

summary: Diffuse intrinsic pontine glioma (DIPG) is an incurable brain tumor that arises in the pons of children.  Recent studies strongly suggest that roughly 30% of DIPG tumors contain copy number increases in platelet derived growth factor receptor A (PDGFRA). We established a novel mouse model by knocking in the human PDGFRA conditional to Cre exposure by crossing in germline Olig2-Cre (specific to oligodendrocytes). This knock in is used in conjuction with p53 tumor suppressor knockout conditional to cre exposure as well. Mutants contain the PDGFRA KI allele, while wild-types only contain p53 floxed alleles with Olig2-Cre. At 18 days of age, mutants demonstrate tremors and hind limb paralysis, and die shortly thereafter. The goal is to determine the differences in brain stems near the time of death in the presence of the PDGFRA mutation when compared to controls without this mutation.

overall_design: Examination of gene expression differences in mouse brainstem of PDGFRA overexpression

first: Herminio

last: Cardona


title: METTL14 facilitates global genome repair and suppresses skin tumorigenesis

city: Chicago

first: Xiaolong

last: Cui

state: IL

address: 680 N Lake Shore Drive

department: Department of Preventive Medicine

institute: Northwestern University

summary: Global genome repair (GGR), a subpathway of nucleotide excision repair (NER), corrects bulky helix-distorting DNA lesions across the whole genome, and is essential for preventing mutagenesis and skin cancer. Here we show that METTL14 (methyltransferase-like 14), a critical component of the m6A RNA methyltransferase complex, promotes GGR through regulating m6A mRNA methylation-mediated DDB2 translation and suppresses UV-induced skin tumorigenesis. Ultraviolet B irradiation (UVB) down-regulates METTL14 protein through NBR1-dependent selective autophagy. METTL14 knockdown decreases GGR and DDB2 abundance. Conversely, overexpression of wild-type METTL14, but not its enzymatically inactive mutant, increases GGR and DDB2 abundance. METTL14 knockdown decreases m6A methylation and translation of the DDB2 transcripts. Adding DDB2 reverses the GGR repair defect in METTL14 knockdown cells, indicating that METTL14 facilitates GGR through regulating DDB2 m6A methylation and translation. Similarly, knockdown of YTHDF1, an m6A reader promoting translation of m6A modified transcripts, decreased DDB2 protein levels. Both METTL14 and YTHDF1 bind to the DDB2 transcript. In mice, skin-specific heterozygous METTL14 deletion increases UV-induced skin tumorigenesis. Furthermore, METTL14 as well as DDB2 is down-regulated in human and mouse skin tumors and by chronic UV irradiation in mouse skin, and METTL14 level is associated with the DDB2 level, suggesting a tumor-suppressive role of METTL14 in UV-associated skin tumorigenesis in association with DDB2 regulation. Taken together, these findings demonstrate that METTL14 is a target for selective autophagy and acts as a critical epitranscriptomic mechanism to regulate GGR and suppresses UVB-induced skin tumorigenesis.

overall_design: m6A IP seq of control, METTL14-knockdown, or UVB-irradiated HaCaT keratinocytes in duplicate

first: Jiangbo

last: Wei

first: Zizhao

last: Yang


title: RNA-sequencing of CD4+ T cells isolated from  C57BL/6 and B6.NZMSle1/Sle2/Sle3 treated with ant-CD45RB

city: Nashville

first: Christopher

middle: Scott

last: Wilson

state: Tennessee

address: 2213 Garland Ave, MRBIV (Langford) 7415

institute: Vanderbilt University Medical Center

summary: C57BL/6 or B6.NZMSle1/Sle2/Sle3 were treated with 100ug of anti-CD45RB (MB23G2) on days 0,1,3,5,and 7. Mice were sacrificied and spleens were harvested and sorted for untouched CD4+ T cells using negative Magentic-activated cell sorting. RNA was isolated and the subjected to RNA sequencing.

overall_design: Examination of the transcriptome in CD4+ T cells in healthy and lupus prone mice in response to a tolerizing immune therapy (anti-CD45RB).

middle: S

first: Daniel

last: Moore


title: TRPS1 drives heterochromatic origin refiring and cancer genome evolution

first: Yu

last: Zhang

address: Xueyuan Road

institute: Peking University Health Science Center

summary: The molecular etiology underlying TRPS1 loss-of-function in tricho-rhino-phalangeal syndrome (TRPS) remains to be understood. Intriguingly, TRPS1 is also bioinformatically postulated as a breast cancer driver. We report here that TRPS1 is prevalently amplified in breast cancer and promotes breast carcinogenesis. We showed that the tumorigenic potential of TRPS1 is derived from its nucleation of the replication machinery and its enforced replication of H3K9me3-marked heterochromatic regions. During early stage of genome duplication, TRPS1 encourages chromatin loading of the anaphase-promoting complex (APC/C) and subsequent degradation of Geminin, an inhibitor for the assembly of the pre-replication complex, leading to uncontrolled refiring of the heterochromatic replication origins. Overexpression of wild-type but not TRPS-associated loss-of-function mutants of TRPS1 is sufficient to drive genome amplification that mimics the genomic alterations in cancer, and TRPS1-evoked aberrant genome dynamically evolves to confer therapeutic resistance to a broad spectrum of treatment compounds. Together, these results link TRPS1 to heterochromatin replication, highlighting the sufficiency of a single oncogene to drive cancer genome evolution.

overall_design: We examined the binding profiles of TRPS1 and its associated factors, the DNA replication patterns in control and TRPS1 knockdown cells, and the extrachromosomal DNA in control and TRPS1 overexpressed cells. siNS = non-specific control siRNA. siTRPS1 = TRPS1 siRNA.

first: Jianguo


title: Integrating whole blood transcriptomic collection procedures into the current anti-doping testing system, including long-term storage and re-testing of anti-doping samples

city: Rome

first: Giscard

middle: Humberto Oliveira

last: Lima

address: Piazza Lauro de Bosis, 15

country: Italy

institute: University of Rome Foro Italico

summary: Recombinant human erythropoietin administration studies involving transcriptomic approaches have demonstrated a gene-expression signature that could aid detection of blood doping. However, current anti-doping testing does not involve blood collection into tubes with RNA preservative. This study investigated if whole blood in long-term storage and whole blood leftover from standard haematological testing in short-term storage could be used for transcriptomic analysis despite lacking RNA preservative. Whole blood samples were collected from thirteen and fourteen healthy  males, for long-term and short-term storage experiments. Long-term storage: whole blood collected into Tempus™ tubes and K2EDTA tubes and subjected to long-term (i.e., −80°C) storage and RNA extracted. After storage, K2EDTA tubes were thawed and extracted using GeneJET RNA Purification Kit (Thermo Fisher Scientific, Vilnius, Lithuania) or Tempus™ Spin RNA Isolation Kit (Life Technologies, Carlsbad, CA, USA). RNA quality and purity was sufficient for gene expression analysis. Principle Component Analysis of microarray and RNA-seq gene expression data for long-term storage: When comparing gene expression between blood tubes with and without RNA preservation, 6% (4058 transcripts) were differentially expressed. RNA quantity, purity and integrity was not significantly compromised from long-term storage in blood storage tubes lacking RNA preservative, indicating that transcriptomic analysis could be conducted using anti-doping samples collected or biobanked without RNA preservation.

overall_design: We analyzed whole blood samples with the highest RNA Integrity Number from 8 male healthy subjects using the Affymetrix Human Exon 1.0 ST platform in the long-term study design. Array data was processed by Affymetrix Exon Array Computational Tool. No techinical replicates were performed.

first: Alexander

last: Kolliari-Turner

first: Fernanda

middle: R

last: Malinsky

first: Fergus

last: Guppy

first: Renan

last: Martin

first: Guan

last: Wang

first: Sven

last: Voss

first: Costas

last: Georgakopoulos

first: Paolo

last: Borrione

first: Fabio

last: Pigozzi

first: Yannis

last: Pitsiladis


title: Krill oil treatment increases distinct PUFAs and oxylipins in adipose tissue and liver and attenuates obesity-associated in-flammation via direct and indirect mechanisms

city: Rijswijk

first: Eveline

last: Gart

state: Zuid-Holland

address: Van vredenburchweg 659

institute: TNO

summary: The development of obesity is characterized by metabolic overload of tissues and sub-sequent organ inflammation. Health effects of krill oil (KrO) on obesity-associated inflammation remain largely elusive, because long-term treatments with KrO have not been performed so far. In this study, mice were fed an obesogenic diet (HFD) containing 3% (w/w) KrO for 28 weeks. KrO treatment increased the concentrations of EPA and DHA and associated oxylipins, including 18-HEPE, RvE2, 14-HDHA in white adipose tissue (WAT) and liver. Simultaneously, KrO decreased arachidonic acid (ARA) concentrations and ARA-derived oxylipins (e.g. HETEs, PGD2, PGE2, PGF2α, TXB2). In epididymal WAT (eWAT), KrO activated regulators of adipogenesis (e.g. PPARγ, CEBPα, KLF15, STAT5A), induced a shift towards smaller adipocytes and increased total adipocyte numbers indicative for hyperplasia. KrO reduced crown-like structures in eWAT, and suppressed HFD-stimulated inflammatory pathways including TNFα and CCL2/MCP-1 signaling. The ob-served changes in eWAT were accompanied by reduced leptin and increased adiponectin levels over time, and an improvement in insulin resistance (HOMA-IR). In liver, KrO suppressed in-flammatory signaling pathways including those controlled by IL-1β and M-CSF, without affecting liver histology. Furthermore, KrO deactivated hepatic REL-A/p65-NF-κB signaling, consistent with increased PPARα protein expression and a trend increase in IkBα. In conclusion, long-term KrO treatment increased several anti-inflammatory PUFAs and oxylipins in WAT and liver. These changes were accompanied by beneficial effects on general metabolism and inflammatory tone at tissue level. Stimulation of adipogenesis by KrO allows for safe fat storage and may, together with more direct PPAR-mediated anti-inflammatory mechanisms, attenuate inflammation.

overall_design: Groups of mice (5-15 mice/group) were treated for 28 weeks in 3 diet groups: Chow (control diet), HFD (High Fat Diet), HFD_KrO (HFD + Krill Oil).

first: Kanita

last: Salic

first: Martine

last: Morrison

first: Martien

last: Caspers

first: Wim

last: van Duyvenvoorde

first: Marieke

last: Heijnk

first: Martin

last: Giera

first: Ivana

last: Bobeldijk-Pastorova

first: Jaap

last: Keijer

first: Andreas

last: Storsve

first: Petter-Arnt

last: Hals

first: Robert

last: Kleemann


title: X-linked recessive TLR7 deficiency in 1% of men under 60 years  with life-threatening COVID-19

status: Public on Aug 11 2021

first: Yelena

last: Nemirovskaya

address: 1230 York Avenue

institute: Rockefeller University

summary: Autosomal inborn errors of type I IFN immunity and autoantibodies against these cytokines underlie at least 10% of critical COVID-19 pneumonia cases. We report very rare, biochemically deleterious X-linked TLR7 variants in 16 unrelated male individuals aged 7 to 71 years (mean: 36.7 years) from a cohort of 1,202 male patients aged 0.5 to 99 years (mean: 52.9 years) with unexplained critical COVID-19 pneumonia. None of the 331 asymptomatically or mildly infected male individuals aged 1.3 to 102 years (mean: 38.7 years) tested carry such TLR7 variants (p = 3.5 x 10-5). The phenotypes of five hemizygous relatives of index cases infected with SARS-CoV-2 include asymptomatic or mild infection (n=2, 5 and 38 years), or moderate (n=1, 5 years), severe (n=1, 27 years), or critical (n=1, 29 years) pneumonia. Two boys (aged 7 and 12 years) from a cohort of 262 male patients with severe COVID-19 pneumonia (mean: 51.0 years) are hemizygous for a deleterious TLR7 variant. The cumulative allele frequency for deleterious TLR7 variants in the male general population is < 6.5x10-4. We also show that blood B-cell lines and myeloid cell subsets from the patients do not respond to TLR7 stimulation, a phenotype rescued by wild-type TLR7. The patients’ blood plasmacytoid dendritic cells (pDCs) produce low levels of type I IFNs in response to SARS-CoV-2. Overall, X-linked recessive TLR7 deficiency is a highly penetrant genetic etiology of critical COVID-19 pneumonia, in about 1.8% of male patients below the age of 60 years. Human TLR7 and pDCs are essential for protective type I IFN immunity against SARS-CoV-2 in the respiratory tract.

overall_design: Type I IFN responses to SARS-CoV-2 infection in TLR7-deficient pDCs

first: Takaki

last: Asano

first: Bertrand

last: Boisson

first: Peng

last: Zhang

first: Jean-Laurent

last: Casanova


title: USP22 overexpression leads to aberrant signal transduction but is not sufficient to drive tumor formation in mice

city: Smithville

first: Evangelia

last: Koutelou

state: TX

address: 1808 Park Road 1C

department: Epigenet & Mol Carcinogenesis

institute: MD Anderson Cancer Center

summary: USP22 overexpression is observed in several human cancers and is correlated with poor patient outcomes.  The molecular basis underlying this correlation is not clear. USP22 is the catalytic subunit of the deubiquitylation module in the SAGA histone modifying complex, which regulates gene transcription.  Our previous work demonstrated that loss of USP22 in mice leads to decreased expression of several components of receptor tyrosine kinase and TGFβ signaling pathways.  To determine whether these pathways are upregulated when USP22 is overexpressed, we created a mouse model that expresses high levels of USP22 in all tissues.  Phenotypic characterization of these mice revealed over branching of the mammary glands in females. Transcriptomic analyses indicate upregulation of key pathways involved in mammary gland branching in mammary epithelial cells derived from the USP22 overexpressing mice, including estrogen receptor, ERK/MAPK and TGFβ signaling.  However, USP22 overexpression did not lead to increased tumorigenesis in any tissue. Our findings indicate that elevated levels of USP22 are not sufficient to induce tumors but it may enhance signaling abnormalities associated with oncogenesis.

overall_design: To determine changes in gene expression profile upon Usp22 ubiquitous overexpression in adult nulliparous females, total RNA from mammary gland epithelial cells  of seven pairs of littermate – matched 3.5-month wild-type and Rosa26KI-Usp22 OE/OE mice was extracted for deep sequencing. Key genes identified from RNAseq were validated by qRT-PCR using RNA from the same samples that were used for sequencing.

first: Xianghong

last: Kuang

first: Sharon

middle: Y

last: Dent


title: Core-shell organization in the mouse claustrum

city: Vancouver

first: Mark

middle: Steven

last: Cembrowski

state: BC

address: 2350 Health Sciences Mall

department: Cellular and Physiological Sciences

country: Canada

institute: University of British Columbia

summary: Excitatory neurons form the long-range outputs of the claustrum. Here, we examined diversity in this cell type using single-cell RNA-seq.

overall_design: Examination of excitatory claustrum neuron transcriptomes.


title: AMULET: A novel read count-based method for effective multiplet detection from single nucleus ATAC-seq data

city: Farmington

first: Duygu

last: Ucar

state: Connecticut

address: 10 Discovery Drive

institute: The Jackson Laboratory

summary: Similar to other droplet-based single cell assays, single nucleus ATAC-seq (snATAC-seq) data harbor multiplets that confound downstream analyses. Detecting multiplets in snATAC-seq data is particularly challenging due to data sparsity and limited dynamic range (0 reads: closed chromatin, 1: open on in one parental chromosome allele, 2: open on in both alleles chromosomes). Yet, these unique data features offer an opportunity to identify multiplets. ATAC-DoubletDetector (https://ucarlab.github.io/ATAC-DoubletDetector/) AMULET (Atac MULtiplet Estimation Tool) exploits these unique features to detect multiplets by studying enumerates the number of regions with >2 uniquely aligned reads across the genome to effectively detect multiplets - an effective alternative to methods based on artificially-generated multiplets. We evaluated the method by generating snATAC-seq data (e.g., state-of-the-art ArchR). For benchmarking we generated snATAC-seq data and generated data fromeasured the efficacy of AMULET inm in two primary human tissues: peripheral human blood mononuclear cells (PBMCs) and pancreatic islet samples. AMULET detects had high multiplets with an estimated precision (estimated via donor-based multiplexing) and high recall (estimated via simulated doublets) compared to alternatives 0.57 precision and achieves 0.85 recall. When and was the most effective when a certain read depth is achieved (a certain read depth per nucleus is achieved samples are sequenced deeply (e.g., median read count per nucleus >20K25K) reads per nucleus in PBMCs), ATAC-DoubletDetector captured 85% of simulated doublets (i.e., recall), significantly outperforming ArchR (24%). For lower read depth, ATAC-DoubletDetector and ArchR produced complementary results. Moreover, ATAC-DoubletDetector was equally effective in identifying homotypic multiplets (i.e., multiplets from the same cell type), which are missed by simulation-based methods. Cell-specific marker peaks enabled accurate (85%) tracing of cellular origins of snATAC-seq multiplets. Accordingly, more abundant cells within a tissue are more likely to form multiplets and the majority of multiplets are homotypic. ATAC-DoubletDetector is a fast and effective multiplet detection/annotation tool for improved single cell epigenomic data analyses across diverse biological systems and conditions.

overall_design: ATAC-seq reads of human pancreatic islet single cells from 1 male and 1 female donor. ATAC-seq reads of PBMC single cells from 3 multiplexed donors.

first: Asa

last: Thibodeau

first: Alper

last: Eroglu

first: Chris

last: McGinnis

first: Nathan

last: Lawlor

first: Romy

last: Kursawe

first: Danny

last: Conrad

first: Zev

last: Gartner

first: Michael

middle: L

last: Stitzel